The role of sperm inner acrosomal membrane proteins in acrosomal biogenesis and fertilization

受精是发育的基石，两个分化的细胞，精子和卵子为胚胎发生和以后的发育奠定了基础，配子的能力是必不可少的。然而，对于导致受精卵发育的精卵相互作用所涉及的分子和机制，人们只有初步的了解。考虑到30- 50%的男性因素不育病例没有已知的原因，这一点得到了强调。我们了解这些机制的策略是分离和分子解剖参与受精过程的精子室（如顶体和核周卵泡）。这些蛋白的特性为研究细胞间室的重要性提供了线索，并允许我们设计特异性探针（如抗体、野生型和突变型重组体以及竞争性抑制剂）来测试这些蛋白在受精和/或精子发生过程中的预测功能和机制。利用这种方法，在NSERC发现基金的授权下，我和我的学生实习生研究了顶体内膜（IAM）的功能。这种精子成分在受精过程中起着至关重要的作用，因为它通过顶体反应暴露出来，然后将精子牢固地结合到卵子的透明带（ZP）上，这是随后由内酰胺引导的精子穿透细胞外基质的必要步骤。在我们的研究之前，IAM的蛋白质是未知的。我们已经鉴定并表征了以下IAM蛋白：IAM38， ZP结合受体；MMP2，电位ZP穿透酶；原丙氨酸，潜在的MMP激活酶；SPACA1，跨膜蛋白，可能组织并将这些蛋白附着在IAM上；SubH2Bv， KAPa和KAPß在精子发生过程中形成复合体并可能引导顶体囊泡到达细胞核。本研究旨在验证这些蛋白的潜在功能并阐明其作用机制。所获得的知识无疑将有助于更好地了解精子不育缺陷的起源及其诊断。它还有助于改进辅助生殖技术的应用和设计一种避孕方法。

Fertilization is the cornerstone of development where the two differentiated cells, sperm and egg set the stage for embryogenesis and beyond, for which competency of the gametes is essential. Yet only rudimentary knowledge is available on the molecules and mechanisms involved in sperm-egg interactions leading to zygotic development. This is underscored in consideration that 30-50 % of male-factor infertility cases have no known cause. Our strategy in gaining an understanding of these mechanisms is to isolate and molecularly dissect sperm compartments (e.g., acrosome and perinuclear theca) that are involved in the fertilization process. The identities of the proteins provides clues to the compartment's significance and allows us to design specific probes (e.g., antibodies, and wild type and mutant recombinants and competitive inhibitors) to test the predicted functions and mechanisms of these proteins during fertilization and/or spermiogenesis. Utilizing this approach, under the mandate of the NSERC Discovery Grant, my student trainees and I have investigated the function of the inner acrosomal membrane (IAM). This sperm component plays a critical role during fertilization as it is exposed by the acrosome-reaction and then binds the sperm firmly to the zona pellucida (ZP) of the egg, a compulsory step for subsequent IAM-directed sperm penetration of this extracellular matrix. Before our study the proteins of the IAM were unexplored. We have identified and characterized the following IAM proteins: IAM38, binding receptor to ZP; MMP2, potential ZP penetrating enzyme; proacrosin, potential MMP activating enzyme; SPACA1, trans-membrane protein which potentially organizes and attaches these proteins to the IAM; SubH2Bv, KAPa and KAPß which form a complex and potentially direct the acrosomic vesicle to the nucleus during spermiogenesis. The present proposal intends to validate the potential functions of these proteins and elucidate their mechanisms of action. The knowledge gained would unquestionably lead to a better understanding of the origins of sperm infertility defects and their diagnosis. It could also serve to improve application of Assisted Reproductive Technologies and to design a contraceptive.